The long term objective of these studies is to elucidate how steroid (estradiol) and protein (FSH and LH) hormones interact to regulate ovarian follicular growth. One aspect of granulosa cell differentiation which is unique and required for follicular ovulation and luteinization is the positive manner in which FSH, along with estradiol, regulates an increase in the FSH receptor-adenylate cyclase-cAMP dependent protein kinase response system. If the mechanisms by which these hormones act to enhance granulosa cell function were to be elucidated, a major contribution to our knowledge of follicular development would be achieved. Accordingly, the specific aims of the proposed research are: 1) to determine the mechanisms by which estradiol and FSH increase FSH responsive adenylate cyclase activity in the absence of changes in FSH receptor content; 2) to determine if the hormonally induced increases in the binding of cAMP to the regulatory subunit of type II cAMP dependent protein kinase (RII) are mediated by cAMP and if they are associated with changes in the synthesis and/or activity of the catalytic subunit (c); and 3) to identify membrane, cytoskeletal and cytosol proteins whose phosphorylation is related to changes in granulosa cell function and luteiniation. To accomplish these aims we intend 1) to examine changes in the amount/activity of individual components of the adenylate cyclase system (FSH receptor, nucleotide binding or N protein and catalytic subunit); 2) to determine the relative changes in the synthesis/degradation of protein kinase (RII, C) and its inhibitor (PKI) and 3) to identify and characterize cAMP and Ca++ dependent phosphorylation of proteins in granulosa cells of follicles at different stages of development. Because granulosa cells go from a relatively cyclase-kinase "deficient" state in preantral follicles to a cyclase-kinase "enriched" state in preovulatory follicles, granulosa cells offer a unique system in which to relate changes in protein phosphorylation with specific physiological events such as receptor induction and receptor loss, increased and decreased cyclase activity, steroidogemesis, and luteinization.